Early life experience influences both hormonal and behavioral responses to stress throughout life in the rat model. Indeed, early-life experiences may modulate human stress-responses and coping, with long-term implications for emotional health and cognitive function. The mechanisms by which neonatal events lead to long-term alterations in the expression and function of stress-related effectors have not been established. Particularly, the nature of the critical, early steps resulting in permanent alteration in glucocorticoid receptors and corticotropin releasing hormone (CRH) expression remain largely unresolved. Work from the applicant's laboratory has demonstrated a rapid, robust modulation of the expression of CRH, a key transducer of the stress-response, in hypothalamus and in amygdala following daily handling and recurrent stress in neonatal rats. This proposal tests the hypothesis that early, rapid and persistent changes of CRH expression in hypothalamus and amygdala play a critical role in the mechanism(s) by which early-life experience influences the hormonal stress-response long-term. Therefore, the proposed experiments will determine whether: (1) alteration in CRH expression in hypothalamus and amygdala is an early result of the handling experience, preceding changes in glucocorticoid receptors, (2) this effect involves the activation of the trans-acting transcription factor, CREB, (3) Handling-induced alterations of stress responses occur concurrently with alterations in CRH-mRNA, and prior to the onset of changes in glucocorticoid receptors. The precise timing and sequence of handling - induced molecular changes, and the transition from temporary to stable alterations in these components of the brain adrenal axis will also be established. The results of the proposed studies will further our understanding of the Neurobiology of Stress. In particular, they will determine the role of CRH in the pivotal, early stages of the molecular cascade by which early life experience permanently alters responses to stress. Thus, these studies will set the stage for early intervention, with a promise of influencing the consequences of early-life experience on neurobiological correlates of stress throughout life.